Disrupted phylogeographical microsatellite and chloroplast DNA patterns indicate a vicariance rather than long‐distance dispersal origin for the disjunct distribution of the Chilean endemic Dioscorea biloba (Dioscoreaceae) around the Atacama Desert

Aim The Chilean endemic Dioscorea biloba (Dioscoreaceae) is a dioecious geophyte that shows a remarkable 600 km north–south disjunction in the peripheral arid area of the Atacama Desert. Its restricted present‐day distribution and probable Neogene origin indicate that its populations have a history linked to that of the Atacama Desert, making this an ideal model species with which to investigate the biogeography of the region. Location Chile, Atacama Desert and peripheral arid area. Methods Two hundred and seventy‐five individuals from nine populations were genotyped for seven nuclear microsatellite loci, and plastid trnL–F and trnT–L sequences were obtained for a representative subset of these. Analyses included the estimation of genetic diversity and population structure through clustering, Bayesian and analysis of molecular variance analyses, and statistical parsimony networks of chloroplast haplotypes. Isolation by distance was tested against alternative dispersal hypotheses. Results Microsatellite markers revealed moderate to high levels of genetic diversity within populations, with those from the southern Limarí Valley showing the highest values and northern populations showing less exclusive alleles. Bayesian analysis of microsatellite data identified three genetic groups that corresponded to geographical ranges. Chloroplast phylogeography revealed no haplotypes shared between northern and southern ranges, and little haplotype sharing between the two neighbouring southern valleys. Dispersal models suggested the presence of extinct hypothetical populations between the southern and northern ranges. Main conclusions Our results are consistent with prolonged isolation of the northern and southern groups, mediated by the life‐history traits of the species. Significant isolation was revealed at both large and moderate distances as gene flow was not evident even between neighbouring valleys. Bayesian analyses of microsatellite and chloroplast haplotype diversity identified the southern area of Limarí as the probable area of origin of the species. Our data do not support recent dispersal of D. biloba from the southern range into Antofagasta, but indicate the fragmentation of an earlier wider range, concomitant with the Pliocene–Pleistocene climatic oscillations, with subsequent extinctions of the Atacama Desert populations and the divergence of the peripheral ones as a consequence of genetic drift.     

An Insertion Peptide in Yeast Glycyl-tRNA Synthetase Facilitates both Productive Docking and Catalysis of Cognate tRNAs

The yeast Saccharomyces cerevisiae possesses two distinct glycyl-tRNA synthetase (GlyRS) genes: GRS1 and GRS2. GRS1 is dually functional, encoding both cytoplasmic and mitochondrial activities, while GRS2 is dysfunctional and not required for growth. The protein products of these two genes, GlyRS1 and GlyRS2, are much alike but are distinguished by an insertion peptide of GlyRS1, which is absent from GlyRS2 and other eukaryotic homologues. We show that deletion or mutation of the insertion peptide modestly impaired the enzyme''s catalytic efficiency in vitro (with a 2- to 3-fold increase in K_m and a 5- to 8-fold decrease in k_cat). Consistently, GRS2 can be conveniently converted to a functional gene via codon optimization, and the insertion peptide is dispensable for protein stability and the rescue activity of GRS1 at 30°C in vivo. A phylogenetic analysis further showed that GRS1 and GRS2 are paralogues that arose from a gene duplication event relatively recently, with GRS1 being the predecessor. These results indicate that GlyRS2 is an active enzyme essentially resembling the insertion peptide-deleted form of GlyRS1. Our study suggests that the insertion peptide represents a novel auxiliary domain, which facilitates both productive docking and catalysis of cognate tRNAs.     

Effect of alkyl side chain variation on the electron-transfer activity of ubiquinone derivatives

The effect of the alkyl side chain of the ubiquinone molecule on the electron-transfer activity of ubiquinone in mitochondrial succinate-cytochrome c reductase is studied by using synthetic ubiquinone derivatives that possess the basic ubiquinone structure of 2,3-dimethoxy-5-methyl-1,4-benzoquinone with different alkyl side chains at the 6-position. The alkyl side chains vary in chain length, degree of saturation, and location of double bonds. When a ubiquinone derivative is used as an electron acceptor for succinate-ubiquinone reductase, an alkyl side chain of six carbons is needed to obtain the maximum activity. However, when it serves as an electron donor for ubiquinol-cytochrome c reductase or as a mediator in succinate-cytochrome c reductase, an alkyl side chain of 10 carbons gives maximal efficiency. Introduction of one or two isolated double bonds into the alkyl side chain of the ubiquinone molecule has little effect on electron-transfer activity. However, a conjugated double bond system in the alkyl side chain drastically reduces electron-transfer efficiency. The effect of the conjugated double bond system on the electron-transferring efficiency of ubiquinone depends on its location in the alkyl side chain. When location is far from the benzoquinone ring, the effect is minimal. These observations together with the results obtained from photoaffinity-labeling studies lead us to conclude that flexibility in the portion of the alkyl side chain immediately adjacent to the benzoquinone ring is required for the electron-transfer activity of ubiquinone.     

ACE/ACE2 Ratio and MMP-9 Activity as Potential Biomarkers in Tuberculous Pleural Effusions

Objective: Pleural effusion is common problem, but the rapid and reliable diagnosis for specific pathogenic effusions are lacking. This study aimed to identify the diagnosis based on clinical variables to differentiate pleural tuberculous exudates from other pleural effusions. We also investigated the role of renin-angiotensin system (RAS) and matrix metalloproteinase (MMPs) in the pathogenesis of pleural exudates.Experimental design: The major components in RAS and extracellular matrix metabolism, including angiotensin converting enzyme (ACE), ACE2, MMP-2 and MMP-9 activities, were measured and compared in the patients with transudative (n = 45) and exudative (n = 80) effusions. The exudative effusions were come from the patients with tuberculosis (n = 20), pneumonia (n = 32), and adenocarcinoma (n = 28).Results: Increased ACE and equivalent ACE2 activities, resulting in a significantly increased ACE/ACE2 ratio in exudates, were detected compared to these values in transudates. MMP-9 activity in exudates was significantly higher than that in transudates. The significant correlation between ACE and ACE2 activity that was found in transudates was not found in exudates. Advanced analyses showed significantly increased ACE and MMP-9 activities, and decreased ACE2 activity in tuberculous pleural effusions compared with those in pneumonia and adenocarcinoma effusions. The results indicate that increased ACE and MMP-9 activities found in the exudates were mainly contributed from a higher level of both enzyme activities in the tuberculous pleural effusions.Conclusion: Interplay between ACE and ACE2, essential functions in the RAS, and abnormal regulation of MMP-9 probably play a pivotal role in the development of exudative effusions. Moreover, the ACE/ACE2 ratio combined with MMP-9 activity in pleural fluid may be potential biomarkers for diagnosing tuberculous pleurisy.     

A high‐throughput method to quantify feeding rates in aquatic organisms: A case study with Daphnia

1. Food ingestion is one of the most basic features of all organisms. However, obtaining precise—and high‐throughput—estimates of feeding rates remains challenging, particularly for small, aquatic herbivores such as zooplankton, snails, and tadpoles. These animals typically consume low volumes of food that are time‐consuming to accurately measure.
2. We extend a standard high‐throughput fluorometry technique, which uses a microplate reader and 96‐well plates, as a practical tool for studies in ecology, evolution, and disease biology. We outline technical and methodological details to optimize quantification of individual feeding rates, improve accuracy, and minimize sampling error.
3. This high‐throughput assay offers several advantages over previous methods, including i) substantially reduced time allotments per sample to facilitate larger, more efficient experiments; ii) technical replicates; and iii) conversion of in vivo measurements to units (mL^‐1 hr^‐1 ind^‐1) which enables broad‐scale comparisons across an array of taxa and studies.
4. To evaluate the accuracy and feasibility of our approach, we use the zooplankton, Daphnia dentifera, as a case study. Our results indicate that this procedure accurately quantifies feeding rates and highlights differences among seven genotypes.
5. The method detailed here has broad applicability to a diverse array of aquatic taxa, their resources, environmental contaminants (e.g., plastics), and infectious agents. We discuss simple extensions to quantify epidemiologically relevant traits, such as pathogen exposure and transmission rates, for infectious agents with oral or trophic transmission.

     

MV-mimicking micelles loaded with PEG-serine-ACP nanoparticles to achieve biomimetic intra/extra fibrillar mineralization of collagen in vitro

Since their discovery, matrix vesicles (MVs) containing minerals have received considerable attention for their role in the mineralization of bone, dentin and calcified cartilage. Additionally, MVs' association with collagen fibrils, which serve as the scaffold for calcification in the organic matrix, has been repeatedly highlighted. The primary purpose of the present study was to establish a MVs–mimicking model (PEG-S-ACP/micelle) in vitro for studying the exact mechanism of MVs-mediated extra/intra fibrillar mineralization of collagen in vivo. In this study, high-concentration serine was used to stabilize the amorphous calcium phosphate (S-ACP), which was subsequently mixed with polyethylene glycol (PEG) to form PEG-S-ACP nanoparticles. The nanoparticles were loaded in the polysorbate 80 micelle through a micelle self-assembly process in an aqueous environment. This MVs–mimicking model is referred to as the PEG-S-ACP/micelle model. By adjusting the pH and surface tension of the PEG-S-ACP/micelle, two forms of minerals (crystalline mineral nodules and ACP nanoparticles) were released to achieve the extrafibrillar and intrafibrillar mineralization, respectively. This in vitro mineralization process reproduced the mineral nodules mediating in vivo extrafibrillar mineralization and provided key insights into a possible mechanism of biomineralization by which in vivo intrafibrillar mineralization could be induced by ACP nanoparticles released from MVs. Also, the PEG-S-ACP/micelle model provides a promising methodology to prepare mineralized collagen scaffolds for repairing bone defects in bone tissue engineering.     

磷酸肌醇激酶FAB1调控拟南芥根毛伸长

FAB1/PIKfyve是介导PI(3,5)P₂ (磷脂酰肌醇3,5-二磷酸)生物合成的磷酸肌醇激酶。在动物和酵母(Saccharomyces cerevisiae)中, PI(3,5)P₂参与调控胞内膜运输, 但在植物中的研究较少。该文通过分析拟南芥(Arabidopsis thaliana) FAB1的T-DNA插入突变体的表型解析PI(3,5)P₂的生物学功能。拟南芥FAB1基因家族包含FAB1A、FAB1B、FAB1C和FAB1D四个基因。研究发现, fab1a/b呈现雄配子体致死的表型。利用遗传杂交获得fab1b/c/d三突变体, 发现FAB1B、FAB1C和FAB1D功能缺失导致根毛相比野生型变短, 经FAB1特异性抑制剂YM201636处理后的野生型中也观察到相似的短根毛表型。此外, fab1b/c/d三突变体中DR5转录水平降低。同时, 外源施加生长素类似物2,4-D和NAA能部分恢复fab1b/c/d植株短根毛的表型, 但fab1b/c/d突变体对生长素转运抑制剂(1-NOA和TIBA)的敏感性与野生型相似。此外, FAB1B/C/D功能缺失使根毛中ROS的含量减少且影响肌动蛋白的表达。上述结果表明, FAB1B/C/D通过调控生长素分布、ROS含量和肌动蛋白的表达影响拟南芥根毛伸长。